Bottom Line:
Proteobacteria were identified as the dominant phylum within both subsurface and thermal spring environments, but only one genera, Rheinheimera, was identified among all samples.Using Morisita similarity indices as a metric for pairwise comparisons between sites, we found that the communities of thermal springs are highly distinct from subsurface datasets.Although the Limpopo thermal springs do not appear to provide a new window for viewing subsurface bacterial communities, we report that the taxonomic compositions of the subsurface sites studied are more similar than previous results would indicate and provide evidence that the microbial communities sampled at depth are more correlated to subsurface conditions than geographical distance.

ABSTRACTSouth Africa has numerous thermal springs that represent topographically driven meteoric water migrating along major fracture zones. The temperature (40-70°C) and pH (8-9) of the thermal springs in the Limpopo Province are very similar to those of the low salinity fracture water encountered in the South African mines at depths ranging from 1.0 to 3.1 km. The major cation and anion composition of these thermal springs are very similar to that of the deep fracture water with the exception of the dissolved inorganic carbon and dissolved O2, both of which are typically higher in the springs than in the deep fracture water. The in situ biological relatedness of such thermal springs and the subsurface fracture fluids that feed them has not previously been evaluated. In this study, we evaluated the microbial diversity of six thermal spring and six subsurface sites in South Africa using high-throughput sequencing of 16S rRNA gene hypervariable regions. Proteobacteria were identified as the dominant phylum within both subsurface and thermal spring environments, but only one genera, Rheinheimera, was identified among all samples. Using Morisita similarity indices as a metric for pairwise comparisons between sites, we found that the communities of thermal springs are highly distinct from subsurface datasets. Although the Limpopo thermal springs do not appear to provide a new window for viewing subsurface bacterial communities, we report that the taxonomic compositions of the subsurface sites studied are more similar than previous results would indicate and provide evidence that the microbial communities sampled at depth are more correlated to subsurface conditions than geographical distance.

Figure 2: Thermal spring taxonomic distribution. A bar plot of the relative abundance (x-axis) of various phyla (color) per thermal spring site (y-axis). Due to its high relative abundance, the phylum proteobacteria was split into its corresponding classes. Members of the “Other” bin include: Armatimonadetes, Spirochaetes, SR1, and Synergisetes.

Mentions:
Rarefaction curves (Supplement Figure 5) revealed that sequences from Eiland, Mphephu, Sagole and Siloam thermal springs captured more of the diversity within those sites than the limited number of sequences from Souting (n = 132) and Tshipise (n = 120) thermal springs did. A comparison of the observed number of genera vs. the number estimated by the Chao1 parameter (Chao, 1984) suggested that the sequences represent 47% (Tshipise) to 68% (Mphephu) of the predicted total number of genera present in the thermal springs (Table 3). All thermal spring samples yielded taxonomic distributions that were found to be significantly different (p < 0.001) from one another through both weighted and unweighted pairwise Unifrac significance tests (Lozupone and Knight, 2005). Four of the six thermal springs (Eiland, Siloam, Souting, and Tshipise) were found to be dominated (>50%) by Proteobacteria, whereas approximately 55% of Mphephu's sequences belonged to Bacteroidetes. Sagole's most abundant phylum was Cyanobacteria (32%) (Figure 2). Souting was found to be the thermal spring site with the highest evenness (Pielou's evenness = 0.83; Table 3) with 30 different genera of Gammaproteobacteria present. Eiland exhibited the lowest Pielou's evenness (0.57) with 60% of sequences belonging to two genera of Proteobacteria: Hydrogenophaga (class Betaproteobacteria) and Stenotrophomonas (class Gammaproteobacteria). Sequences from Siloam contained the highest number of unique genera (119), whereas Tshipise contained the fewest unique genera (30) (Table 3). No sequences derived from Tshipise were related to Alphaproteobacteria.

Figure 2: Thermal spring taxonomic distribution. A bar plot of the relative abundance (x-axis) of various phyla (color) per thermal spring site (y-axis). Due to its high relative abundance, the phylum proteobacteria was split into its corresponding classes. Members of the “Other” bin include: Armatimonadetes, Spirochaetes, SR1, and Synergisetes.

Mentions:
Rarefaction curves (Supplement Figure 5) revealed that sequences from Eiland, Mphephu, Sagole and Siloam thermal springs captured more of the diversity within those sites than the limited number of sequences from Souting (n = 132) and Tshipise (n = 120) thermal springs did. A comparison of the observed number of genera vs. the number estimated by the Chao1 parameter (Chao, 1984) suggested that the sequences represent 47% (Tshipise) to 68% (Mphephu) of the predicted total number of genera present in the thermal springs (Table 3). All thermal spring samples yielded taxonomic distributions that were found to be significantly different (p < 0.001) from one another through both weighted and unweighted pairwise Unifrac significance tests (Lozupone and Knight, 2005). Four of the six thermal springs (Eiland, Siloam, Souting, and Tshipise) were found to be dominated (>50%) by Proteobacteria, whereas approximately 55% of Mphephu's sequences belonged to Bacteroidetes. Sagole's most abundant phylum was Cyanobacteria (32%) (Figure 2). Souting was found to be the thermal spring site with the highest evenness (Pielou's evenness = 0.83; Table 3) with 30 different genera of Gammaproteobacteria present. Eiland exhibited the lowest Pielou's evenness (0.57) with 60% of sequences belonging to two genera of Proteobacteria: Hydrogenophaga (class Betaproteobacteria) and Stenotrophomonas (class Gammaproteobacteria). Sequences from Siloam contained the highest number of unique genera (119), whereas Tshipise contained the fewest unique genera (30) (Table 3). No sequences derived from Tshipise were related to Alphaproteobacteria.

Bottom Line:
Proteobacteria were identified as the dominant phylum within both subsurface and thermal spring environments, but only one genera, Rheinheimera, was identified among all samples.Using Morisita similarity indices as a metric for pairwise comparisons between sites, we found that the communities of thermal springs are highly distinct from subsurface datasets.Although the Limpopo thermal springs do not appear to provide a new window for viewing subsurface bacterial communities, we report that the taxonomic compositions of the subsurface sites studied are more similar than previous results would indicate and provide evidence that the microbial communities sampled at depth are more correlated to subsurface conditions than geographical distance.

ABSTRACTSouth Africa has numerous thermal springs that represent topographically driven meteoric water migrating along major fracture zones. The temperature (40-70°C) and pH (8-9) of the thermal springs in the Limpopo Province are very similar to those of the low salinity fracture water encountered in the South African mines at depths ranging from 1.0 to 3.1 km. The major cation and anion composition of these thermal springs are very similar to that of the deep fracture water with the exception of the dissolved inorganic carbon and dissolved O2, both of which are typically higher in the springs than in the deep fracture water. The in situ biological relatedness of such thermal springs and the subsurface fracture fluids that feed them has not previously been evaluated. In this study, we evaluated the microbial diversity of six thermal spring and six subsurface sites in South Africa using high-throughput sequencing of 16S rRNA gene hypervariable regions. Proteobacteria were identified as the dominant phylum within both subsurface and thermal spring environments, but only one genera, Rheinheimera, was identified among all samples. Using Morisita similarity indices as a metric for pairwise comparisons between sites, we found that the communities of thermal springs are highly distinct from subsurface datasets. Although the Limpopo thermal springs do not appear to provide a new window for viewing subsurface bacterial communities, we report that the taxonomic compositions of the subsurface sites studied are more similar than previous results would indicate and provide evidence that the microbial communities sampled at depth are more correlated to subsurface conditions than geographical distance.